The present invention relates generally to treatment of disease in humans and other mammals. More particularly, it relates to biological markers such as cell populations, cell surface antigen expression levels, and soluble factor concentrations that are indicative of the efficacy of treatment of atopic asthma and other inflammatory disorders.
Prednisone is a corticosteroid used to treat a wide variety of inflammatory disorders, including asthma, atopy, arthritis, multiple sclerosis, ulcerative colitis, and Crohn""s disease. While a comprehensive understanding of the action of prednisone and other glucocorticoids (a class of corticosteroids) is lacking, the drugs are known to have broad-ranging anti-inflammatory and immunosuppressive effects, including inhibition of pro-inflammatory mediators and activation of anti-inflammatory mediators. They affect the growth, differentiation, and function of monocytes and lymphocytes; the distribution of cellular subsets; and the production of cytokines, cellular proteins that are secreted and affect the behavior of other cells. Because the disorders treated by glucocorticoids themselves involve unknown immunological mechanisms, it is unclear which of the many effects of prednisone are most important in inhibiting the inflammatory response.
In addition, because of their broad-ranging systemic effects, prednisone and other glucocorticoids have a large number of side effects, some of them quite serious, which restrict their applicability in many patients, particularly for long-term use. These side effects include weight gain, hyperglycemia, bone thinning, digestive problems, cataracts, susceptibility to infection, hypertension, mood swings, and insomnia. It would be beneficial to have a drug that could provide the desired anti-inflammatory and immunosuppressive effects of prednisone without the detrimental side effects. Additionally, localizing the effects to the region of inflammation (e.g., the lungs) would minimize the systemic side effects. In order to do so, however, more information must be obtained about the mechanism of action of glucocorticoids in treating inflammatory disorders.
One very common inflammatory disease that has long been treated with prednisone is asthma, a chronic respiratory syndrome of uncertain etiology. Typical asthma symptoms include coughing, wheezing, chest tightness, and shortness of breath. These clinical symptoms are thought to result from hyper-responsiveness of the airways and a long-term inflammatory process that causes reversible obstruction of the airways. Many asthma sufferers also suffer from atopy, a hypersensitive allergic response to airborne antigens. The clinical manifestations of atopic asthma arise from the superposition of environmental factors on genetic predispositions that increase the probability of developing the syndrome.
Atopic asthma is an immunologic disease mediated by IgE antibodies and characterized by infiltration of the airways with mast cells, lymphocytes, and eosinophils.
The allergic response results from a hyperactivity of TH2 (type 2 helper) T lymphocytes, triggering the production of cytokines such as interleukin (IL)-4, IL-5, IL-6, IL-10, and IL-13, which enhance antibody production from B cells and induce essential aspects of the allergic response, such as mucosal tissue injury by eosinophils. T cells also support IgE-mediated responses to airborne allergens and orchestrate the recruitment and activation of primary effector cells. Products released by the inflammatory cells accumulated in the airways contribute to the tissue destruction characteristic of asthma.
A large number of studies have been performed, mostly in vitro, to elucidate the anti-inflammatory mechanism of glucocorticoids in diseases such as asthma. One hypothesis is that glucocorticoids suppress the activation of cells that produce cytokines to prime eosinophils and induce the migration of lymphocytes, eosinophils, and basophils into the airway. Upon entering a cell, glucocorticoids bind with intracellular glucocorticoid receptors (GR), which are widely distributed among different cell types. The glucocorticoid-receptor complex then enters the cell nucleus and turns on specific genes by binding with DNA and directing the transcription process. In particular, glucocorticoids interact with two transcription factors, activating protein 1 (AP-1) and nuclear factor NF-xcexaB. AP-1 is involved in the regulation of several genes, including those that express adhesion molecules and cytokines, while NF-xcexaB regulates the transcription of genes involved in the inflammatory response. It is estimated that each cell type has ten or more target genes per cell, although they may not all be expressed. It would be highly desirable to develop new glucocorticoids or other drugs that would be more selective moderators of gene expression, leading to a reduction in toxic effects and localization of effects to disease regions.
Current studies on the effect of glucocorticoids on asthma and other inflammatory disorders are performed by measuring the effect of the drug on quantities of cell types, cell surface antigen expression, and soluble factors, both in vivo and in vitro. A number of factors have been shown to be correlated with glucocorticoid use, such as an increase in circulating granulocytes or decrease in number of eosinophils, which are responsible for many of the inflammatory tissue damage effects of asthma. However, while many studies examine IgE levels and related factors, there are few data available with respect to other measurements in vivo, such as cell surface marker expression on granulocytes and lymphocytes, or soluble factors in serum. Available data are also conflicting. For example, variable effects of glucocorticoids on T cell counts have been shown, while B cells are believed to be only minimally affected by glucocorticoids, with redistribution from peripheral blood to other lymphoid compartments being of main importance. There have also been conflicting reports on immunoglobulin levels post glucocorticoid treatment. As is well known to those of skill in the art, the correlation between in vitro and in vivo measurements is tenuous at best, and in vivo measurements must be performed to obtain information that can be used for treatment or diagnostic purposes.
Methods and compositions are currently being developed for alternative treatments of asthma and other inflammatory disorders that minimize side effects. Much of this work is devoted to regulation of the various interleukins. For example, U.S. Pat. No. 5,908,839, issued to Levitt et al., discloses methods for treating asthma by regulating the function of the IL-9 receptor. U.S. Pat. No. 5,683,983, issued to Barrett et al., discloses compounds that bind to the IL-5 receptor. U.S. Pat. No. 5,874,080, issued to Hebert et al., provides anti-IL-8 monoclonal antibodies for treatment of asthma.
In general, much more information must be obtained before less broad ranging but sufficiently effective anti-inflammatory drugs can be developed for treatment of autoimmune or inflammatory diseases such as asthma. In addition, accurate but simple methods for evaluating drug efficacy are lacking. One of the problems in elucidating both drug action and disease pathogenesis is that appropriate tools are lacking to measure a broad range of applicable immunological components in vivo from a large number of subjects. Thus many studies on the effects of glucocorticoids on cellular responses are performed in vitro or in animal models, neither of which is directly applicable to humans. In addition, the studies tend to examine factors that are already known to be implicated in glucocorticoid action, rather than searching for novel factors that may be useful for indicating the disease progression or treatment efficacy.
There is a need, therefore, for a simple but effective method of gauging the anti-inflammatory or immunosuppressive response of candidate drugs for treating asthma, atopy, and other inflammatory diseases. There is also a need for more information about the mechanism by which glucocorticoids inhibit the inflammatory response. Further, there is a need for methods to determine systemic versus local effects of administered drugs.
The present invention addresses these needs by providing novel biological markers (biomarkers) indicative of anti-inflammatory or immunosuppressive action of a drug in a subject. These novel biomarkers can be used to assess the anti-inflammatory action of a drug, detect systemic versus local effects of an anti-inflammatory drug, and better understand the mechanism of action of glucocorticoids such as prednisone. The information obtained can aid in selection of an appropriate glucocorticoid, dose, and administration route. Additionally, the information provided by the present invention can help in designing next generation drugs that have strong anti-inflammatory effects but fewer or less significant side effects than existing glucocorticoids.
The inventive biomarkers include cell populations, cell surface antigens, and soluble factors whose measurement values in a biological sample change significantly (either increase or decrease) after an anti-inflammatory drug is administered to the subject from whom the sample is obtained. In particular, these biomarkers include CD89 expression on granulocytes, CD38 expression on CD4 T cells, HLA class II expression on B cells, CD62L expression on B cells, monocyte count, HLA class II expression on monocytes, MMP-3 concentration, and SAA concentration.
In one embodiment, the invention provides a method for determining whether a candidate drug is effective in treating an inflammatory or autoimmune disease. Specific doses and delivery routes can also be examined. The method is performed by administering the candidate drug to a subject; obtaining a biological sample, such as a blood sample, from the subject; measuring the level of at least one of the inventive biological markers in the biological sample; and comparing the measured level with a standard level. Typically, the standard level is obtained by measuring the same marker or markers in the subject before drug administration. Depending upon the difference between the measured and standard levels, the drug can be considered to have an anti-inflammatory or immunosuppressive effect. Typical anti-inflammatory diseases treated by the candidate drug include atopy and asthma. If multiple biomarkers are measured, at least one and up to all of the biomarkers must change significantly, in the expected direction, for the drug to be considered anti-inflammatory. Preferably, multiple markers must change for the drug to be considered effective.
In an alternative embodiment, the present invention provides a method for detecting a systemic effect of an anti-inflammatory drug (e.g., a glucocorticoid such as prednisone) in a subject. In this method, a biological sample correlated with systemic activity, such as blood or urine, is obtained from the subject. Next, a set of factors including at least one of the inventive biomarkers is measured in the biological sample. The measured values are then compared with standard values, preferably measurements of the same biomarkers taken from the same subject before the drug was administered. Preferably, measurements are also made of a local biological sample, one correlated with local rather than systemic activity, extracted from the same subject. The change in local values after drug can be analyzed to determine whether the drug has local effects in addition to or instead of systemic effects.